Investigating the effect of molecular crowding agents on protein stability in a bacterial proteome

ABSTRACT: Molecular crowding refers to the restriction of space available for molecular mobility in solution due to the presence of other macromolecules. Crowding can influence biological phenomena such as protein stability and folding, diffusion, enzyme kinetics, molecular interactions, and phase separation. In contrast to many biochemical studies that employ dilute solutions comprising a small number of reactants, the cellular environment is considered to be highly crowded, with up to 40% of cell volume occupied by many thousands of proteins and other large biomolecules. We examined the protein stability effects of widely used cosolvents that are considered to act as molecular crowders for experimental purposes in a bacterial proteome (Cupriavidus necator) using the Thermal Proteome Profiling (TPP) method. While all six tested compounds (Ficoll 70, Ficoll 400, dextran 40, dextran 86, PEG 1, PEG 8) reduced the global mean melting temperature, significantly increased or decreased conformational stability was observed among distinct subsets of the proteome, with some individual proteins showing sensitivity to multiple crowding agents. Proteins displaying enhanced stability in the presence of crowding agents tended to be more hydrophobic, were more likely to show classic enzyme-like properties, and propensity for protein interactions. Our data support a direct binding/preferential exclusion model of enhanced stability, rather than alternative models based on viscosity or crowding.

INSTRUMENT(S):

ORGANISM(S): Cupriavidus Necator (strain Atcc 17699 / H16 / Dsm 428 / Stanier 337) (ralstonia Eutropha) Bacteria

SUBMITTER: Eugene Dillon  

LAB HEAD: Gerard Cagney

PROVIDER: PXD067043 | Pride | 2026-02-23

REPOSITORIES: Pride